This proposal represents a continuation of our studies on the development and function of HLA-A2.1 restricted CTL in transgenic murine lines. Two different types of transgenic lines have been developed. One expresses the intact A2.1 molecules, whereas a second line expresses a chimeric form of A2 consisting of the alpha1+alpha2 domains of A2 linked to the alpha3 domain of mouse Kb. Previous studies demonstrated A2-restricted responses were obtained more readily from A2/Kb transgenics, due to enhanced interaction of the A2/Kb molecule with mouse CD8. When A2- restricted, influenza specific responses were obtained from A2 transgenics they demonstrated cross-reactive recognition of A2/Kb targets in the absence of specific antigen. We have proposed this as a consequence of positive selection of a T cell repertoire by a class I molecule that interacts weakly with CD8. To compensate for the suboptimal contribution by CD8 to the overall avidity of the T cell for the A2 restriction molecule, the affinity of the TCR must be higher than normal to achieve positive selection. The combination of the usually high TCR affinity for the restriction element, plus the enhanced affinity of interaction between CD8 and A2/Kb, results in recognition of A2/Kb in the absence of cognate antigen. This proposal will test this hypothesis, as well as the possibility that the endogenous peptide recognized in association with A2/Kb may represent the same peptide responsible for positive selection of that particular TCR in the A2-expressing thymus. To this end, we will construct TCR transgenics expressing such a receptor and study the MHC and peptide requirements for its positive and negative selection, both in vivo and in vitro. Previous studies have demonstrated that A2-restricted responses in human and mice recognize the same endogenously processed epitope peptides. Thus, transgenic mice expressing functional HLA molecules represent an excellent model system for human T cell recognition. As a second aim of this proposal, we will use the A2/Kb transgenic line as a model for evaluating the utility of A2 restricted responses specific for the human p53 tumor-suppressor gene in eliminating tumor cells. These studies should develop a prototype for in vivo evaluation of A2 restricted responses to well defined viral and tumor specific proteins of potential value as targets for immunotherapy and vaccine design.